螢光發報之原位適配接合反應應用於異戊二烯焦磷酸合成酶抑制劑的開發

張皓晴

doi:10.6342/NTU.2009.01738

3.239.109.55，Hello！

Close
Search Quick Search Advanced Search Publication Search	Personal Service Member Login Searching History Use Bonus Card SYMSKAN
Browse Browse by University of Graduate Thesis Browse by Pinyin of Graduate Thesis Browse by Conference Proceedings Topic Browse by Conference Proceedings Publisher Browse by Pinyin of Conference Proceedings Browse by eBook Category Browse by eBook Topic Browse by eBook Publisher
Customer Services Authorize Airiti Recommend a Journal

Search Symbol (Half-width)	Description of Search Symbols
Space	"AND" indicates the intertwining of key terms used in a search
Double Quotation Marks ("") ( " " )	Double quotation marks indicate the beginning and end of a phrase, and the search will only include terms that appear in the same order of those within the quotations. Example: "image process" ： " image process "
?	Indicates a variable letter. Entering two ? will indicate two variable letters, and so on. Example: "Appl?", search results will yield apple, apply… e , appl y … ( (often used to English word searches) )
*	Indicates an unlimited number of variable letters to follow, from 1~n. Example: Enter "appl*", search results will yield apple, apples, apply, applied, application…(often used in English word searches) e , appl es , appl y , appl ied , appl ication … ( (often used to English word searches) )
AND、OR、NOT	Boolean logic combinations of key words is a skill used to expand or refine search parameters. (1) AND (1) AND: Refines search parameters (2) OR (2) OR: Expands search parameters (3) NOT: Excludes irrelevant parameters

DOI stands for Digital Object Identifier （ D igital O bject I dentifier ） ,
and is the unique identifier for objects on the internet. It can be used to create persistent link and to cite articles.

Using DOI as a persistent link

To create a persistent link, add「http://dx.doi.org/」「 http://dx.doi.org/ 」 before a DOI.
For instance, if the DOI of an article is 10.5297/ser.1201.002 , you can link persistently to the article by entering the following link in your browser: http://dx.doi.org/ 10.5297/ser.1201.002 。
The DOI link will always direct you to the most updated article page no matter how the publisher changes the document's position, avoiding errors when engaging in important research.

Cite a document with DOI

When citing references, you should also cite the DOI if the article has one. If your citation guideline does not include DOIs, you may cite the DOI link.

DOIs allow accurate citations, improve academic contents connections, and allow users to gain better experience across different platforms. Currently, there are more than 70 million DOIs registered for academic contents. If you want to understand more about DOI, please visit airiti DOI Registration （ doi.airiti.com ）。

Data Source

Basic & Applied Sciences > Chemistry
College of Science > Graduate Institute of Chemistry

Bibliography Management Tool

Related Links

Report an Issue

Purchase Single Articles

Download PDF

螢光發報之原位適配接合反應應用於異戊二烯焦磷酸合成酶抑制劑的開發

Fluorogenic In Situ Click Chemistry Applied to Inhbitor Discovery of Prenyl Pyrophosphate Synthases

張皓晴 , Masters　　Advisor：陳昭岑　　

繁體中文 DOI： 10.6342/NTU.2009.01738

螢光；原位適配接合化學；異戊二烯焦磷酸；抑制劑；篩選； fluorogenic ； in situ click ； prenyl pyrophosphate ； inhibitor ； screening

Abstract │ Reference (75) │ Altmetrics

Abstract 〈TOP〉

Parallel Abstract 〈TOP〉

Isoprenoids are widespread in the biological systems. For examples, abnormal cellular signal transduction via protein prenylation with C20-GGPP is observed in some tumor cells, and the C55-UPP serves as a lipid carrier in the bacterial cell wall biosynthesis. Therefore, developing inhibitors for prenyl pyrophosphate synthases has great implications in the biomedical applications. In this thesis, we have developed an effective screening method based on the fluorogenic in situ click reactions to fish out inhibitors for a series of prenyl pyrophosphate synthases more effecitvely.
     We designed and synthesized several hydrophobic 3-azidocoumarins and hydrophilic arylalkynes, to complement the hydrophobic tunnel and the polar catalytic region of the target enzymes. Utilizing the strategy of in-situ target-guided synthesis, we are hoping that the enzymes select the proper fragments themselves and further promote the azide-alkyne cycloaddition between the hydrophobic fragments having 3-azidocoumarin and the hydrophilic ones with alkyne functionality. Once the cycloadduct formed, the fluorescence of azicocoumarin is no longer suppressed by the azido group and the resultant fluorescence enhancement is a good indication that the target enzyme promotes the click reaction itself.
     The binding ability of 10 hydrophobic fragments to the active sites of UPPs was evaluated by fluorescence competition experiments by first incubating the enzyme with MANT-GPP, which was proven to be the substrate and inhibitor for UPPs and could give fluorescence increase at 425 nm once bounded. The titration results revealed that the binding ability of the hydrophobic fragments could attributed to the length of substituted hydrocarbon tails. Hence, we only employed three different lengths of prenyl substituent (denoted as Pre-hAzC, Ger-hAzC, Far-hAzC) and three hydrophilic arylalkynes (denoted as pPP, mPP, mBP) with potent pyrophosphate or bisphosphonate groups were chosen for further in situ click experiments. These fragments (3 x 3) were arranged in a spatial combinatorial fashion and each combination was incubated with three target enzymes (UPPs, OPPs or hGGPPs) for 3 days, respectively. The fluorescence was then measured by the fluorescence plate reader for each combination. These results showed that the combination of Pre-hAzC/mBP incubated with hGGPPs gave the best fluorescence with a 4.5-fold fluorescence enhancement among 27 (3 x 3 x 3) possible combinations. To confirm and verify which is the isomer of Pre-hAzC/mBP generated by the in situ click reactions, the possible cycloadducts of Pre-hAzC and mBP, denoted as Pre-1,4-mBP and Pre-1,5-mBP, were synthesized and tested their biological activity. The inhibition results revealed that the former suppresses 84% of hGGPPs activity at 100 uM concentration, the latter only inhibits 15% of hGGPPs activity. Moreover, the molecular modeling results of docking are consistent with the biological inhibition data.

Reference ( 75 ) 〈TOP〉

1. Keserü, G. M.; Makara, G. M. Nat. Rev. Drug Discov. 2009, 8, 203-212.
連結：
2. Lipinski, C.; Hopkins, A. Nature 2004, 432, 855-861.
連結：
3. Rideout, D. Science 1986, 233, 561-563.
連結：
4. Rideout, D. Cancer Invest. 1994, 12, 189-202.
連結：
5. Maly, D. J.; Choong, I. C.; Ellman, J. A. Proc. Natl. Acad. Sci. USA 2000, 97, 2419-2424.
連結：

Altmetrics

〈TOP〉

E-mail ：

When an article is available to download, a notice will be sent to your mailbox address.

E-mail ：